Recently, matrix-assisted laser desorption ionization (MALDI) has been gaining acceptance as a way to ionize analytes for time of flight (TOF) mass spectrometry (MS), especially for analysis of large molecules such as proteins. Generally, a matrix material (e.g., 2,5-dihydroxybenzoic acid, nicotinic acid, cinnamic acid derivatives such as sinapinic acid) and analytes are applied to a surface so that crystals of the matrix materials and analyte solids are formed thereon. Typically, laser radiation is used for ionization of the analytes. After ionization, the ions can be separated (and therefore identified) according to their mass-to-charge ratios in mass spectrometry.
In analyzing large molecules, such as proteins, peptides, and nucleic acids, gel electrophoresis (GE) is a routinely used technique. MS is considered to be very useful for identifying analytes separated in GE. For interfacing gel electrophoresis with MS, membranes have been evaluated as sample supports for MALDI-MS (e.g., Blackledge et al., Anal. Chem., 1995, 67, 843-848; Zaluzec et al., J. Am. Soc. Mass. Spectrom., 1994, 5, 230-237; Vesting et al., 1994, Anal. Chem., 66. 471-477). To take advantage of this technology, analytical samples are placed on a repeller in the mass spectrometer to be ionized and repelled. However, some analytical samples are not amenable to be readily used for MALDI-MS. The process of transferring analytical samples from other analytical apparatuses to MALDI-MS can be labor-intensive (e.g., transferring protein from a gel to a membrane by electroblotting). Even worse, the samples may not contain a high enough concentration of analytes.
Capillary electrophoresis (CE) has also emerged as a powerful tool for efficient separation of analytes in an aqueous solution. CE analysis is conducted in a capillary that is immersed in an electrolyte. When an analytical sample is introduced into the capillary and an electrical potential is applied to the two ends of the capillary, the resulting electric field causes analytes to migrate along the capillary, as well as draws the electrolyte through the capillary. The analytes move from one end of the capillary to the other end, each at a rate dependent on its electrophoretic mobility and the rate of fluid flow in the capillary. To collect samples of analytes exiting a CE system for MALDI-MS, the fractions exiting the CE capillary need to be collected and applied on a substrate that is suitable for supporting the ionization of analytes in the MS.
Carson et al. (U.S. Pat. No. 5,126,025, also BioTechniques, 1993, 14(1):51-55) describes an apparatus for collecting fractions from capillary electrophoresis on a membrane. The apparatus is described as having a capillary tube, a porous layer containing an electrolyte, a first electrode adjacent the entrance end of the capillary, and a second electrode in electrical contact with the porous layer. A liquid absorbent layer or a containerful of liquid is in contact with the porous layer to supply it with the electrolyte (to wet the porous layer). There is no indication that this system can be used to collect samples for MALDI-MS analysis. The electrolyte (in constant contact with the porous layer and being supplied continuously by the absorbent or container) will cause diffusion and spreading of the analytes exiting the capillary on the porous layer, thereby lowering the concentration of analytes and resulting in analyte-containing spots or streaks with large surface areas. This will result in less accuracy if used in MALDI-MS analysis of the unknown sample. What is needed is an apparatus and method for collecting fractions from samples eluting from a capillary in CE to result in analytecontaining spots of higher concentrations and smaller surface areas.